Interpretive Summary: In this investigation, icMS profiling has been used to discriminate between serum samples collected from control rats and rats exposed to dietary TM for 12 months based on the presence of CDT. The icMS approach was found to be very specific with transferrin and IgG representing the largest peaks in the mass spectra and produced a promising classification model. It should be noted that although a single protein was targeted here, icMS can be multiplexed for the simultaneous enrichment of several proteins that could be used to build ion signatures based on multiple peaks (Wang et al., 2008) thereby potentially enhancing the accuracy of the model. Such additional candidate biomarkers of TM exposure could include a1-microglobulin, a1-antitrypsin and kininogen 1, which have been identified previously (Penno et al., 2009a). While the practicality of an icMS-based diagnostic assay for disease detection is presently limited by the availability of suitable MS instruments in the clinical laboratory, the increasing popularity of microbial identification by MALDI-TOF MS (Seng et al., 2010) is expected to bring MS-based diagnostics into mainstream clinical usage. Accordingly, the development of this assay represents a significant step towards a high-throughput screening test for detecting CT exposure that may be suitable for both livestock and humans.

Technical Abstract:
The neurological livestock disease annual ryegrass toxicity (ARGT) is caused by the ingestion of the naturally occurring glycolipid toxins – the corynetoxins. Corynetoxins also threaten human health as potential contaminants of the food supply. Presently, there are no routine diagnostic tests for corynetoxins-exposure in humans or livestock. Chronic ingestion of corynetoxins has been modeled in rats exposed to dietary tunicamycins for 12 months and carbohydrate deficient transferrin (CDT) has been previously identified as a candidate disease biomarker. Here, the technique of immuno-capture mass spectrometry (icMS) was used to evaluate serum levels of CDT, discriminating between control and tunicamycins-exposed rats with 85% accuracy. The icMS approach is based on the combination of specific transferrin enrichment with functionalized magnetic beads and automated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). With no other clinically-relevant diagnostic tests available icMS could be readily adapted for high-throughput clinical assessment of corynetoxins-exposure in humans or livestock.